Humulus yunnanensis plant named ‘Kriya’

ABSTRACT

A new humulus plant named ‘Kriya’ is disclosed. The leaves of this new humulus plant contain a cannabinoid level of ˜15-23 mg/g and the inflorescence of this new humulus plant contain a cannabionoid level of ˜124-142 mg/g.

FIELD OF THE INVENTION

The present invention relates to a new and distinct Humulus plant. Theplant is botanically known as Humulus yunnanensis var kriya.

BACKGROUND OF THE INVENTION

The new and distinct humulus plant originated from a cross hybridizationof feral H. yunnanensis variants collected from the Pekong area withinthe Arunachal Pradesh region of India. Various H. yunnanensis sampleswere collected for analysis from various regions of India, including thegroves in Puging, Singing, and Pekong, as well as in Mouling NationalPark, Kaying, and Lipo. H. yunnanensis male and female saplings withroots were collected, along with male and female flowers. All collectedsamples were tested for the presence of cannabinoids using standardmethods known in the art (see Korte. F. and Sieper. H., J. Chromatoqr.13:90 (1964), which is hereby incorporated by reference in itsentirety). Only 5.2% of the H. yunnanensis samples collected haddetectable levels of cannabinoids. The average cannabinoid level in theinflorescence of the H. yunnanensis plants containing cannabinoids was2.1 mg/g¹. The cannabinoid level in plant tissue as described throughoutthis application is provided as milligrams of cannabinoid per gram offreeze dried plant material.

The Pekong strains of H. yunnanensis were identified as having unusuallyhigh cannabinoid content, with detectable levels of cannabigerol (CBG),cannabichromene (CBC), cannabidiol (CBD), cannabielsoin (CBE) andcannabidivarin (CBDV) found. The content of cannabidiol, cannabichromeneand cannabigerol was high, usually >85-90% of the carboxylatedcannabinoids and >65-70% of the uncarboxylated cannabinoids. No traceamounts of tetrahydrocannabinol were detected in the Pekong strains.

Table 1 (below) summarizes the inflorescence size and cannabinoid levelof six of the H. yunnanensis plants collected from the Pekong region.The cannabinoid levels are reported as milligram cannabinoid per gram offreeze-dried plant tissue. Of these samples, samples 3, 4, and 6 wereselected for breeding based on their high cannabinoid content. All ofthese samples were negative for the presence of tetrahydrocannabinol.

TABLE 1 Characteristics of Pekong H. yunnanensis Pekong #1 Pekong #2Pekong #3 Inflorescence 3.7 cm 4.8 cm 7.4 cm size (length in cm)Inflorescence 22 mg/g 26 mg/g 56 mg/g Cannabinoid (mg/g) Leaf 6.3 mg/g6.3 mg/g 7.5 mg/g Cannabinoid (mg/g) Pekong #4 Pekong #5 Pekong #6Inflorescence 6.2 cm 5.9 cm 6.6 cm size (length in cm) Inflorescence 41mg/g 32 mg/g 42 mg/g Cannabinoid (mg/g) Leaf 5.3 mg/g 6.1 mg/g 4.8 mg/gCannabinoid (mg/g)

To initiate the generation of ‘Kriya’, Pekong #3 plant was crossed withPekong #6 plant to produce 128 female progeny. The cannabinoid level infemale inflorescence of each plant was assessed. Of the 128 progeny, 74of the plants did not contain a significant cannabinoid level.Twenty-three of the progeny had longer inflorescence (>6 cm), but thelevel of cannabinoid in the inflorescence was less than 20 milligramsper gram of freeze dried tissue. Twenty-four of the progeny had mediuminflorescence (between 4 and 6 cm in length) and a medium inflorescencecannabinoid level (between 25 and 35 mg/g). Seven of the offspring hadinflorescence greater than 7 cm in length and a cannabinoid levelgreater than 70 mg/g. From this analysis it was surmised that thepresence of cannabinoids is a recessive trait in Humulus yunnanensis.

In parallel with the cross of Pekong #3 and Pekong #6, Pekong #3 wasindependently crossed with Pekong #4 plant to generate 128 progeny. Thecannabinoid level in the female inflorescence was also assessed in theseprogeny. Eight plants having an inflorescence cannabinoid level between66 mg/g and 73 mg/g were identified.

First generation plants produced by the cross between Pekong #3 andPekong #6 and the cross between Pekong #3 and Pekong #4 having thehighest cannabinoid level were selected for crossbreeding to producesecond generation plants (n=7 plants from the Pekong #3/Pekong #6 cross;n=8 plants from the Pekong #3/Pekong #4 cross). The offspring of thesecrosses were bred to produce third and fourth generation offspring. Thefifth generation yielded female plants having an average cannabinoidcontent of 128 milligrams cannabinoid per gram of freeze driedinflorescence and 16 milligrams cannabinoid per gram of freeze driedleaves and “trims”. Of these female plants, one plant was chosen forasexual propagation. This new variety of high cannabinoid content plantwas named Humulus yunnanensis var kriya or ‘Kriya’.

Further propagation of female ‘Kriya’ was carried out using in-vitroculture starting on Mar. 14, 2017 in Nainital, Uttarakhand, India. Thehypocotyl and the newly germinating buds of the female ‘Kriya’ plantwere micro-propagated. Sterile plant tissues were removed from an intactplant. A small portion of plant tissue was placed on B5 medium to halfits ionic strength (B5/2). The medium was thickened with agar to createa gel which supported the explant during growth.

The tissue samples produced during the first stage were multiplied toincrease overall number. The tissue was grown into small “plantlets”.Offshoot production was induced by hormone treatment. All propagules of‘Kriya’ have been observed to be true to type in that during all asexualmultiplication, the inflorescences and globose of the original planthave been maintained. After the formation of multiple shoots, theseshoots were transferred to rooting medium with a high auxin\cytokininratio.

The ‘Kriya’ plantlets were transferred to the soil, with vermicompost,from the plant media. After a few days the plantlets were “hardened” andtransferred to the field to grow to full maturity.

BRIEF DESCRIPTIONS OF THE DRAWINGS

FIG. 1 is a photograph of the ‘Kriya’ plant growing in a greenhouse inOoty, India in July 2017.

FIGS. 2A-2B show the Humulus yunnanensis var kriya leaf. FIG. 2A is aphotograph of a seven-lobed ‘Kriya’ leaf and FIG. 2B is a drawing of afive-lobed ‘Kriya’ leaf.

FIG. 3 is a drawing of a young Humulus yunnanensis var kriya femaleinflorescence.

FIGS. 4A-4B are photographs of the Humulus yunnanensis var kriyainflorescence at different stages. FIG. 4A is a picture of a ‘Kriya’inflorescence during the mid-vegetative growth state. FIG. 4B is apicture of a ‘Kriya’ inflorescence during the late reproductive stage.

DETAILED BOTANICAL DESCRIPTION

The present invention relates to a new and distinct humulus plant. Theplant is botanically known as Humulus yunnanensis var kriya.

The following description of ‘Kriya’, unless otherwise noted, is basedon observations made during June through November 2017. Thesemeasurements and ratings were taken from ‘Kriya’ plants planted in March2017.

COMPARATIVE CANNABINOID CHARACTERISTICS

‘Kriya’ is distinguishable from its originating parents and otherrelated varieties of Humulus yunnanensis based on the cannabinoidcontent in its leaves and inflorescence. Table 2 below details thenumber and type of H. yunnanensis samples collected from various regionsin and around India, and the number of these samples that containeddetectable levels of cannabinoids. As indicated in Table 2, many of theH. yunnanensis samples contained no detectable level of cannabinoids,i.e., H. yunnanensis plants collected from Ooty, Puging, Singing,Kaying, and Bomdeling did not contain detectable levels of cannabinoids.

TABLE 2 Comparison of Cannabinoid Content in Samples of H. yunnanensisCollected From India and Bhutan Ooty¹ Samples with Puging² SamplesCannabinoids Samples Samples collected (“CB”) collected w/CB Flowers 190 23 0 Shoot tips 11 0 18 0 Leaves 23 0 36 0 Stem 15 0 11 0 Bark 9 0 8 0Roots 6 0 3 0 Total Collected 83 99 Samples w/C8 0 0 Singing³ Pekong⁴Samples Samples Samples Samples collected w/CB collected w/CB Flowers 410 69 14 Shoot tips 32 0 36 8 Leaves 67 0 103 13 Stem 24 0 43 0 Bark 20 029 0 Roots 8 0 15 0 Total Collected 192 300 Samples w/C8 0 35 Mouling⁵Kaying⁶ Samples Samples Samples Samples collected w/CB collected w/CBFlowers 31 4 29 0 Shoot tips 23 2 21 0 Leaves 47 2 35 0 Stem 12 0 11 0Bark 8 0 6 0 Roots 0 0 4 0 Total Collected 121 106 Samples w/C8 8 0Lipo⁷ Bomdeling⁸ Samples Samples Samples Samples collected w/CBcollected w/CB Flowers 19 4 18 0 Shoot tips 14 2 14 0 Leaves 35 6 33 0Stem 18 0 14 0 Bark 11 0 8 0 Roots 8 0 8 0 Total Collected 105 100Samples w/C8 12 0 Nanda Devi⁹ Samples Samples collected w/CB % with CBFlowers 11 2 Shoot tips 15 2 Leaves 25 2 Stem 8 0 Bark 7 0 Roots 2 0Total Collected 68 1174 Samples w/C8 6 61 5.20% Ooty¹: Ooty village inTamil Nadu, Southern India Puging²: Puging village in Upper Siangdistrict of Arunachal Pradesh India Singing³: Singing village in UpperSiang district of Arunachal Pradesh India Pekong⁴: Pekong village inUpper Siang district of Arunachal Pradesh India Mouling⁵: MoulingNational Park in Arunachal Pradesh India Kaying⁶: Kaying village WestSiang district of Arunachal Pradesh India Lipo⁷: Lipo village West Siangdistrict of Arunachal Pradesh India Bomdeling⁸: Bomdeling WildlifeSanctuary in Bhutan Nanda Devi⁹: Nanda Devi National Park in HimachalPradesh, India

The presence of cannabinoids was most frequently detected in samplescollected from the Pekong region. Table 1 above shows the cannabinoidlevels in the six Pekong plants identified as having the highestcannabinoid levels, including the originating parent plants of ‘Kriya’.Table 3 below shows average cannabinoid content in the inflorescence andleaves of first, second, third, and fourth generation² offspring ofcrossed Pekong samples, as well the average cannabinoid content in theinflorescence and leaves of ‘Kriya’. The average cannabinoid level in‘Kriya’ inflorescence is 133.5 mg/g±8.62 mg/g. This level is >2-foldhigher than the inflorescence cannabinoid level of the original Pekongparental variants (41 mg/g-56 mg/g). This level is also significantlygreater than the average inflorescence cannabinoid level found in first,second, third, and fourth generation plants. First generation plants areprogeny of the cross between Pekong #3 and Pekong #6 plants, and theprogeny of Pekong #3 and Pekong #4 plants. Second generation plants areprogeny of Pekong #3/Pekong #6 offspring crossed with Pekong #3/Pekong#4 offspring. Third generation plants are progeny of second generationcrosses, and fourth generation plants are progeny of third generationcrosses.

The average cannabinoid leaf content of ‘Kriya’ is 19.28±3.75 mg/g. Thisis also >2-fold higher than the leaf cannabinoid content of theoriginating Pekong parental variants (4.8 mg/g-7.5 mg/g). This level isalso significantly greater than the average leaf cannabinoid level foundin the first, second, third, and fourth generation plants.

Cannabidiol (CBD), cannabichromene (CBC), and cannabigerol (CBG) makeup >98% of the cannabinoids present in the inflorescence and leaves of‘Kriya’. Trace amounts (<2%) of cannabielsoin (CBE) and cannbidivarin(CBDV) are also present. As with the parent strains, notetrahydrocannabinol is present in ‘Kriya’.

TABLE 3 Characteristics of ‘Kriya’ and its Predecessors InflorescenceLength Mean Standard Plant N (cm) Deviation Variance 1st Generation 765.767 1.325 1.755 2nd Generation 62 5.933 1.294 1.675 3rd Generation 476.15 1.319 1.739 4th Generation 52 6.783 0.861 0.742 Humulus kriya 368.233 0.857 0.735 Inflorescence Cannabinoid Mean Standard Plant N (mg/g)Deviation Variance 1st Generation 76 33.167 8.06 64.967 2nd Generation62 34.667 8.335 69.467 3rd Generation 47 46.334 7.23 52.267 4thGeneration 52 78.333 8.123 66.967 Humulus kriya 36 133.5 8.62 74.3 LeafCannabinoid Mean Standard Plant N (mg/g) Deviation Variance 1stGeneration 83 1.78 0.342 0.117 2nd Generation 73 2.24 0.623 0.388 3rdGeneration 48 3.4 0.678 0.46 4th Generation 51 6.48 1.105 1.222 Humuluskriya 46 19.28 3.749 14.057

‘Kriya’ is also distinguishable from its originating parental plantsbased on average β-caryophyllene content in the inflorescence andleaves. The average β-caryophyllene level in ‘Kriya’ inflorescence is53.11 mg/g±7.73 mg/g (milligrams of β-caryophyllene per gram offreeze-dried plant tissue). This level is >4-fold higher than theinflorescence β-caryophyllene level of the original Pekong parentalvariants (3 mg/g-11 mg/g). As shown in Table 4 below, the level ofβ-caryophyllene is also markedly different than the averageinflorescence β-caryophyllene level found in first, second, third, andfourth generation plants.

The average β-caryophyllene leaf content of ‘Kriya’ is 10.63±1.99 mg/g.This is almost 10-fold higher than the leaf β-caryophyllene content ofthe originating Pekong parental variants (0.8 mg/g-1.6 mg/g). This levelis also markedly different from the average leaf β-caryophyllene levelfound in the first, second, third, and fourth generation plants.

TABLE 4 β-Caryophyllene Content of ‘Kriya’ and its PredecessorsInflorescence Beta Caryophyllene Mean Standard N (mg/g) DeviationVariance 1st Generation 76 26.77 6.41 41.11 2nd Generation 54 28.52 4.8023.06 3rd Generation 63 36.52 7.81 61.08 4th Generation 47 42.42 8.3369.54 Humulus kriya 73 53.11 7.73 59.82 Leaf Beta Caryophyllene MeanStandard N (mg/g) Deviation Variance 1st Generation 67 2.09 0.411 0.1682nd Generation 73 2.77 0.52 0.28 3rd Generation 59 4.17 0.54 0.29 4thGeneration 71 7.9 1.87 3.49 Humulus kriya 82 10.63 1.99 3.98

‘Kriya’ is also distinguishable from its originating parental plantsbased on average inflorescence size. As indicated in Table 3 above, theaverage length of the inflorescence of ‘Kriya’ is 8.233±0.875 cm, whichis distinctly longer than the size of the inflorescence of theoriginating parent plants (range 6.2 cm-7.4 cm).

BOTANICAL DESCRIPTION

The following botanical description of the new humulus cultivar willvary somewhat depending upon cultural practices and climatic conditions,and can vary with location and season. Quantified measurements areexpressed as an average of measurements taken from a number ofindividual ‘Kriya’ plants. The measurements of any individual plant orany group of plants of the new cultivar may vary from the statedaverage. The color chart used was The Royal Horticulutural SocietyColour Chart (5^(th) ed. 2007).

‘Kriya’ is a perennial vine that is 4-8 feet long, branchingoccasionally (see FIG. 1). The twining habit of ‘Kriya's’ stems allowthis vine to climb adjacent vegetation and fences. The rather stoutstems are light green and longitudinally ridged. Along the ridges ofeach stem, there are rows of stiff prickly hairs. A pair of oppositeleaves occur at intervals along each stem.

The plant has dormant stage (December-February) where the plant shedsall its leaves and only the dried pods remain. The dormant stage isfollowed by a growth stage (February-May), a vegetative stage(June-August), and flowering stage (August-October), after which theplant starts gradually going dormant again. As referred to herein, a“young” plant refers to a plant in the early growth stage (i.e.,February to May), and a “mature” plant refers to a plant at the lateflowering stage (i.e., August to October).

‘Kriya's’ leaves are opposite and cordite, and can grow up to 6 incheslong and 6 inches wide. The leaves are palmate, generally divided into 5lobes with toothed margins; however leaves containing 7 and 9 lobes havebeen observed. Upper leaves are normally 3 lobed and develop to five toseven lobes as they mature. Each lobe is oblanceolate or elliptic inshape with coarsely serrated margins. The upper surface of each leaf ismedium green and moderately covered with short rough hairs (subglabrousand membranous). The lower surface has stiff prickly hairs along themajor veins.

‘Kriya’ normally grows to cover large areas of open ground. It overgrowsshrubs and small trees and can choke them off. It grows rapidly insummer and forms dense foliage several feet deep. It twines aroundshrubs and trees causing them wither. It can displace native vegetation.It is a bushy vine that can be trained to be conical. It takes a conicalappearance when it climbs a trellis or a small tree.

The normal adaxial surface of the leaves are normally subglabrous andmembranous. The normal leaf isn't blistered or curled. During a humidand rainy growing season and cases of “Downy Mildew” caused byPseudoperonospora humuli have been observed. The infected leaves becomebrittle and have downward curling leaves. A yellow color affects theinfected leaves and the spores then appear as purple to black blotcheson the abaxial (underside) of infected leaves.

The rather stout petioles of ‘Kriya’ are as long as, or a little shorterthan, the leaves. These petioles are light green and covered with stiffdownward pointing prickly hairs.

The young female ‘Kriya’ inflorescence is a short spike of flowers withbracts and bracteoles that are ovate-orbiculate. This spike becomesglobose and large with age and tends to nod downward, spanning about5-10 cm long. Flowers originate as pistillate bracts in the leaf axils.The appearance of each spike is dominated by several overlappingpistillate bracts. At the base of each bract, there is a pair ofinconspicuous female flowers. Initially, the pistillate bracts arenarrowly deltoid in shape, but enlarge in size to 6-9 mm long and becomedeltoid with recurved tips and are pilose. These bracts are light todark green, hairy, and strongly ciliate along their smooth margins.These bracts turn a yellowish brown when mature (see FIG. 4). Thebracteoles are more prominent in the young plant, and do not have aflower at the base. Each female flower has a divided style and aninconspicuous calyx that surrounds the developing ovary; there are nopetals (see FIG. 4).

The mature female inflorescences are leafy cone-like catkins orstrobiles. As noted above, when fully developed, the strobiles are about5-10 cm long, oblong in shape and rounded, consisting of a number ofoverlapping, green bracts, attached to a separate axis. If these leafybracts are removed, the axis will be seen to be hairy and to have azigzag path (see FIG. 4). Each of the bracts enfolds at the base a smallfruit (achene). The fruit is an ovoid achene which is about ¼ inch indiameter, roundish, rough, light brown and speckled. The fruit and bracthave translucent glands, which secrete a waxy substance. ‘Kriya’ doesnot secrete the powdery Lupulin which gives the “hops” plant (Humuluslupulus) its distinctive flavor.

-   Disease susceptibility: Aphids, spidermites, powdery mildew, downy    mildew, black root rot, hop mosaic virus, and apple mosaic virus.-   Aroma: Woody aroma, soft complex spice, mild cananga/cinnamon.-   Use: Medicinal.-   Time of flowering: Single flowering season; Autumn.-   Leaves:    -   -   Leaf arrangement.—Opposite.        -   Immaure leaf shape.—Cordate.        -   Mature leaf shape.—Usually a palmate shape, lobed (5-7            lobes). Sometimes simple leaf, also lobed.        -   Leaf apex shape.—Acutely acuminate.        -   Leaf base shape.—Cordate.        -   Abaxial surface.—Rigid spinulose hairs on veins.        -   Abaxial surface color.—140D young plant; 140A mature plant.        -   Adaxial surface.—Densely pubescent.        -   Adaxial surface color.—129D young plant; 130C mature plant.        -   Margin.—Serrate.        -   Average length.—5-15 cm.        -   Average width.—4-15 cm.        -   Cannabinoid content.—15-23 mg/g.-   Petiole:    -   -   Average petiole length.—4-10 cm.        -   Average diameter.—5 mm.        -   Average circumference.—2 cm.        -   Color.—185A.-   Inflorescence: Leafy cone-like catkins or strobiles.    -   -   Arrangement.—Spicate.        -   Shape.—Ovaloid.        -   Average length.—5-10 cm.        -   Average width.—2-4 cm.        -   Strobiles.—Color 153D, 153C, 153B, 153A, 152D, 152C, 152B.        -   Bracts and bracteole.—Subglabrous and membraneous; abaxial            surface has prominent veins. shape: initially narrowly            deltoid; with maturation become enlarged and deltoid with            recurved tips. size: 1.5 cm to 3 cm. color: immature bract            is light to dark green 140D, 140C, 140B, 140A, mature bract            is yellowish brown, 162D, 162C, 162B, 162A. bract apex            length: 0.05-0.80 inch.        -   Cones per node at the side shoot.—Middle of the plant: 1-3            cones. Upper third portion of the plant: 0-1 cones.        -   Total cones per side shoot.—Middle of the plant: 4-16 cones.            Upper third portion of the plant: 0-7 cones.        -   Total cones per plant.—220-840.        -   Achene.—Size: 3 to 9 cm. Shape: flat. Color: 200D.        -   Cannabinoid content.—124-142 mg/g.        -   Flowers.—Color: 149D, 149C.        -   Main shoot anthocyanin coloration.—144C, 144B.

What is claimed:
 1. A new and distinct humulus plant as illustrated anddescribed herein.